Conversion of human cardiac progenitor cells into cardiac pacemaker-like cells.
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ABSTRACT:
SUBMITTER: Raghunathan S
PROVIDER: S-EPMC7035977 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Lineage conversion of one somatic cell type to another is an attractive approach for generating specific human cell types. Lineage conversion can be direct, in the absence of proliferation and multipotent progenitor generation, or indirect, by the generation of expandable multipotent progenitor states. We report the development of a reprogramming methodology in which cells transition through a plastic intermediate state, induced by brief exposure to reprogramming factors, followed by differentiation. We use this approach to convert human fibroblasts to mesodermal progenitor cells, including by non-integrative approaches. These progenitor cells demonstrated bipotent differentiation potential and could generate endothelial and smooth muscle lineages. Differentiated endothelial cells exhibited neo-angiogenesis and anastomosis in vivo. This methodology for indirect lineage conversion to angioblast-like cells adds to the armamentarium of reprogramming approaches aimed at the study and treatment of ischemic pathologies.
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Project description:Self-powered implantable medical electronic devices that harvest biomechanical energy from cardiac motion, respiratory movement and blood flow are part of a paradigm shift that is on the horizon. Here, we demonstrate a fully implanted symbiotic pacemaker based on an implantable triboelectric nanogenerator, which achieves energy harvesting and storage as well as cardiac pacing on a large-animal scale. The symbiotic pacemaker successfully corrects sinus arrhythmia and prevents deterioration. The open circuit voltage of an implantable triboelectric nanogenerator reaches up to 65.2 V. The energy harvested from each cardiac motion cycle is 0.495 μJ, which is higher than the required endocardial pacing threshold energy (0.377 μJ). Implantable triboelectric nanogenerators for implantable medical devices offer advantages of excellent output performance, high power density, and good durability, and are expected to find application in fields of treatment and diagnosis as in vivo symbiotic bioelectronics.
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Project description:BACKGROUND:Human induced pluripotent stem cells (hiPSC) harbor the potential to differentiate into diverse cardiac cell types. Previous experimental efforts were primarily directed at the generation of hiPSC-derived cells with ventricular cardiomyocyte characteristics. Aiming at a straightforward approach for pacemaker cell modeling and replacement, we sought to selectively differentiate cells with nodal-type properties. METHODS:hiPSC were differentiated into spontaneously beating clusters by co-culturing with visceral endoderm-like cells in a serum-free medium. Subsequent culturing in a specified fetal bovine serum (FBS)-enriched cell medium produced a pacemaker-type phenotype that was studied in detail using quantitative real-time polymerase chain reaction (qRT-PCR), immunocytochemistry, and patch-clamp electrophysiology. Further investigations comprised pharmacological stimulations and co-culturing with neonatal cardiomyocytes. RESULTS:hiPSC co-cultured in a serum-free medium with the visceral endoderm-like cell line END-2 produced spontaneously beating clusters after 10-12 days of culture. The pacemaker-specific genes HCN4, TBX3, and TBX18 were abundantly expressed at this early developmental stage, while levels of sarcomeric gene products remained low. We observed that working-type cardiomyogenic differentiation can be suppressed by transfer of early clusters into a FBS-enriched cell medium immediately after beating onset. After 6 weeks under these conditions, sinoatrial node (SAN) hallmark genes remained at high levels, while working-type myocardial transcripts (NKX2.5, TBX5) were low. Clusters were characterized by regular activity and robust beating rates (70-90 beats/min) and were triggered by spontaneous Ca2+ transients recapitulating calcium clock properties of genuine pacemaker cells. They were responsive to adrenergic/cholinergic stimulation and able to pace neonatal rat ventricular myocytes in co-culture experiments. Action potential (AP) measurements of cells individualized from clusters exhibited nodal-type (63.4%) and atrial-type (36.6%) AP morphologies, while ventricular AP configurations were not observed. CONCLUSION:We provide a novel culture media-based, transgene-free approach for targeted generation of hiPSC-derived pacemaker-type cells that grow in clusters and offer the potential for disease modeling, drug testing, and individualized cell-based replacement therapy of the SAN.
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Project description:Glial progenitor cells are widely distributed in brain parenchyma and represent a suitable target for future therapeutic interventions that generate new neurons via in situ reprogramming. Previous studies have shown successful reprogramming of mouse glia into neurons whereas the conversion of human glial cells remains challenging due to the limited accessibility of human brain tissue. Here, we have used a recently developed stem cell-based model of human glia progenitor cells (hGPCs) for direct neural reprogramming by overexpressing a set of transcription factors involved in GABAergic interneuron fate specification. GABAergic interneurons play a key role in balancing excitatory and inhibitory neural circuitry in the brain and loss or dysfunction of these have been implicated in several neurological disorders such as epilepsy, schizophrenia, and autism. Our results demonstrate that hGPCs successfully convert into functional induced neurons with postsynaptic activity within a month. The induced neurons have properties of GABAergic neurons, express subtype-specific interneuron markers (e.g. parvalbumin) and exhibit a complex neuronal morphology with extensive dendritic trees. The possibility of inducing GABAergic interneurons from a renewable in vitro hGPC system could provide a foundation for the development of therapies for interneuron pathologies.
| S-EPMC7698048 | biostudies-literature
Project description:Myocardial function is enhanced by adoptive transfer of human cardiac progenitor cells (hCPCs) into a pathologically challenged heart. However, advanced age, comorbidities, and myocardial injury in patients with heart failure constrain the proliferation, survival, and regenerative capacity of hCPCs. Rejuvenation of senescent hCPCs will improve the outcome of regenerative therapy for a substantial patient population possessing functionally impaired stem cells.Reverse phenotypic and functional senescence of hCPCs by ex vivo modification with Pim-1.C-kit-positive hCPCs were isolated from heart biopsy samples of patients undergoing left ventricular assist device implantation. Growth kinetics, telomere lengths, and expression of cell cycle regulators showed significant variation between hCPC isolated from multiple patients. Telomere length was significantly decreased in hCPC with slow-growth kinetics concomitant with decreased proliferation and upregulation of senescent markers compared with hCPC with fast-growth kinetics. Desirable youthful characteristics were conferred on hCPCs by genetic modification using Pim-1 kinase, including increases in proliferation, telomere length, survival, and decreased expression of senescence markers.Senescence characteristics of hCPCs are ameliorated by Pim-1 kinase resulting in rejuvenation of phenotypic and functional properties. hCPCs show improved cellular properties resulting from Pim-1 modification, but benefits were more pronounced in hCPC with slow-growth kinetics relative to hCPC with fast-growth kinetics. With the majority of patients with heart failure presenting advanced age, infirmity, and impaired regenerative capacity, the use of Pim-1 modification should be incorporated into cell-based therapeutic approaches to broaden inclusion criteria and address limitations associated with the senescent phenotype of aged hCPC.
| S-EPMC3999968 | biostudies-literature
Project description:The heartbeat originates within the sinoatrial node (SAN), a small structure containing <10,000 genuine pacemaker cells. If the SAN fails, the ?5 billion working cardiomyocytes downstream of it become quiescent, leading to circulatory collapse in the absence of electronic pacemaker therapy. Here we demonstrate conversion of rodent cardiomyocytes to SAN cells in vitro and in vivo by expression of Tbx18, a gene critical for early SAN specification. Within days of in vivo Tbx18 transduction, 9.2% of transduced, ventricular cardiomyocytes develop spontaneous electrical firing physiologically indistinguishable from that of SAN cells, along with morphological and epigenetic features characteristic of SAN cells. In vivo, focal Tbx18 gene transfer in the guinea-pig ventricle yields ectopic pacemaker activity, correcting a bradycardic disease phenotype. Myocytes transduced in vivo acquire the cardinal tapering morphology and physiological automaticity of native SAN pacemaker cells. The creation of induced SAN pacemaker (iSAN) cells opens new prospects for bioengineered pacemakers.
| S-EPMC3775583 | biostudies-literature
Project description:In the mammalian heart a conduction system of nodes and conducting cells generates and transduces the electrical signals evoking myocardial contractions. Specialized pacemaker cells initiating and controlling cardiac contraction rhythmicity are localized in an anatomically identifiable structure of myocardial origin, the sinus node. We previously showed that in mammalian embryos sinus node cells originate from cardiac progenitors expressing the transcription factors T-box transcription factor 3 (Tbx3) and Islet-1 (Isl1). Although cardiac development and function are strikingly conserved amongst animal classes, in lower vertebrates neither structural nor molecular distinguishable components of a conduction system have been identified, questioning its evolutionary origin. Here we show that zebrafish embryos lacking the LIM/homeodomain-containing transcription factor Isl1 display heart rate defects related to pacemaker dysfunction. Moreover, 3D reconstructions of gene expression patterns in the embryonic and adult zebrafish heart led us to uncover a previously unidentified, Isl1-positive and Tbx2b-positive region in the myocardium at the junction of the sinus venosus and atrium. Through their long interconnecting cellular protrusions the identified Isl1-positive cells form a ring-shaped structure. In vivo labeling of the Isl1-positive cells by transgenic technology allowed their isolation and electrophysiological characterization, revealing their unique pacemaker activity. In conclusion we demonstrate that Isl1-expressing cells, organized as a ring-shaped structure around the venous pole, hold the pacemaker function in the adult zebrafish heart. We have thereby identified an evolutionary conserved, structural and molecular distinguishable component of the cardiac conduction system in a lower vertebrate.
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Project description:Cardiac repair following MI relies on a finely regulated immune response involving sequential recruitment of monocytes to the injured tissue. Monocyte-derived cells are also critical for tissue homeostasis and healing process. Our previous findings demonstrated the interaction of T and natural killer cells with allogeneic human cardiac-derived stem/progenitor cells (hCPC) and suggested their beneficial effect in the context of cardiac repair. Therefore, we investigated here whether monocytes and their descendants could be also modulated by allogeneic hCPC toward a repair/anti-inflammatory phenotype. Through experimental in vitro assays, we assessed the impact of allogeneic hCPC on the recruitment, functions and differentiation of monocytes. We found that allogeneic hCPC at steady state or under inflammatory conditions can incite CCL-2/CCR2-dependent recruitment of circulating CD14+CD16- monocytes and fine-tune their activation toward an anti-inflammatory profile. Allogeneic hCPC also promoted CD14+CD16- monocyte polarization into anti-inflammatory/immune-regulatory macrophages with high phagocytic capacity and IL10 secretion. Moreover, hCPC bended the differentiation of CD14+CD16- monocytes to dendritic cells (DCs) toward anti-inflammatory macrophage-like features and impaired their antigen-presenting function in favor of immune-modulation. Collectively, our results demonstrate that allogeneic hCPC could reshape monocytes, macrophages as well as DCs responses by favoring their anti-inflammatory/tolerogenic activation/polarization. Thereby, therapeutic allogeneic hCPC might also contribute to post-infarct myocardial healing by modeling the activities of monocytes and their derived descendants.
| S-EPMC5662627 | biostudies-literature
Project description:Cardiac fibroblasts play a key role in chronic heart failure. The conversion from cardiac fibroblast to myofibroblast as a result of cardiac injury, will lead to excessive matrix deposition and a perpetuation of pro-fibrotic signaling. Cardiac cell therapy for chronic heart failure may be able to target fibroblast behavior in a paracrine fashion. However, no reliable human fibrotic tissue model exists to evaluate this potential effect of cardiac cell therapy. Using a gelatin methacryloyl hydrogel and human fetal cardiac fibroblasts (hfCF), we created a 3D in vitro model of human cardiac fibrosis. This model was used to study the possibility to modulate cellular fibrotic responses. Our approach demonstrated paracrine inhibitory effects of cardiac progenitor cells (CPC) on both cardiac fibroblast activation and collagen synthesis in vitro and revealed that continuous cross-talk between hfCF and CPC seems to be indispensable for the observed anti-fibrotic effect.
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